Our brains are packed with trillions of biological transistors known as synapses, which control the flow of information that produces our movements, perceptions, thoughts, and memories. To maintain order and plasticity within this vast interconnected network, neurons must have the ability to turn ON or OFF selected synapses. This project uses the genetic strengths of the model organism C. elegans to investigate the underlying signaling pathways and membrane trafficking mechanisms that allow neurons to regulate signaling across synapses. The pathways of a Ga signaling network control synaptic activity to produce the C. elegans locomotion behavior. These pathways are conserved in all neurons in all animals; however, how they control synaptic activity is not yet understood. This knowledge gap impedes progress in understanding the fundamental mechanisms that drive behavior and human brain functions ranging from sleep to complex thoughts and memories. A guiding hypothesis of this proposal, shaped by new preliminary data, is that the Ga pathways exert their major effects through Dense Core Vesicle (DCV) functions, and that there are important differences in how the pathways regulate DCVs in neuronal cell somas versus axons. Aim 1 of this proposal will use high resolution imaging techniques in living animals to investigate the relative contributions and interactions of the Gaq and Gas pathways in driving neuropeptide release from both cell somas and axons. We will also investigate the broad hypothesis, suggested by our recent studies of DCV maturation, that there is a DCV function unrelated to neuropeptides that is lost when the DCV maturation pathway malfunctions. By defining the maturation pathway in Aims 2 and 3 we hope to obtain clues about this missing function and bring a comprehensive model organism strategy to this under-investigated branch of membrane trafficking. We recently completed a genetic screen in C. elegans for DCV maturation mutants. The screen identified three proteins that, along with UNC-108 (Rab2), are core components in the DCV maturation pathway. These proteins, two of which have undefined functions, are conserved in all organisms with a nervous system, thus highlighting both the fundamental importance and novelty of this pathway. Aim 3 will combine a biochemical approach with a novel genetic approach to add further mechanistic insights to this new pathway. By combining the in vivo relevance of genetics, live animal imaging, and behavioral strategies with the mechanistic relevance of biochemistry, this project has the potential to transform intriguing preliminary findings into fundamental new insights that will begin bridging the gaps between the Ga signaling network, dense core vesicle functions, and behaviors. PUBLIC HEALTH RELEVANCE: This project will investigate nerve cell (neuron) function and signaling in the model organism C. elegans. The 300 neurons in the C. elegans nervous system share many basic functions with the 100 billion neurons in the human nervous system; however, the genes responsible for encoding some of the most critical central neuron functions remain undiscovered or under-investigated, thus impeding progress in understanding and treating human neurological disorders such as schizophrenia, sleep disorders, depression, and learning/ memory disorders. We will use sophisticated genetic experiments in C. elegans - experiments that would be impossible to perform in humans or vertebrate lab animals - to discover and investigate the genes encoding these core functions of neurons.